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Toxicity to microorganisms

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Reference
Endpoint:
activated sludge respiration inhibition testing
Type of information:
experimental study
Adequacy of study:
key study
Study period:
20th August to 1st September 1999
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: The study was conducted in a GLP compliant laboratory according to accepted regulatory guidelines
Qualifier:
according to
Guideline:
OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 850.6800 (Modified Activated Sludge, Respiration Inhibition Test for Sparingly Soluble Chemicals)
Deviations:
no
Qualifier:
according to
Guideline:
other: EEC Commission Directive 87/302/EEC
Deviations:
no
GLP compliance:
yes (incl. certificate)
Details on sampling:
- Concentrations: each flask was sampled
- Sampling method: an aliquot taken from the flask
- Sample storage conditions before analysis: samples analysed without storage
Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
For the purpose of the definitive study the test material was prepared by direct dispersion in water.
Preliminary solubility work conducted showed that due to the extremely viscous nature of the test material, it was impossible to ensure that after dispensing the required amount of test material into a weighing receptacle, the test material could be completely washed into the test vessel. Even the use of ultrasonication did not completely remove the test material from the sides of the weighing receptacle. Therefore in order to ensure that an accurate weight of test material was added to the test vessel, the test material was weighed onto a glass microscope slide prior to addition of the microscope slide and test material to the test vessel.
An amount of test material (500 mgl was weighed onto a glass microscope slide which was dispersed in approximately 250 ml of water and subjected to ultrasonication for approximately 30 minutes. Synthetic sewage (16 ml), activated sewage sludge (200 ml) and water were added to a final volume of 500 ml to give the test concentration of 1000 mg/1.
Test organisms (species):
activated sludge of a predominantly domestic sewage
Details on inoculum:
- Laboratory culture:
- Method of cultivation: continuous aeration at 21C at pH7.5
- Preparation of inoculum for exposure:At time "0" 16 ml of synthetic sewage was diluted to 300 ml with water and 200 ml of inoculum added in a 500 ml conical flask (first control). The mixture was aerated with compressed air via narrow bore glass tubes at a rate of approximately 0.5 - 1 litre per minute.
- Pretreatment: none, innoculum used on day of collection
- Initial biomass concentration: 3.9 g/L suspended solids prior to use
Test type:
static
Water media type:
freshwater
Limit test:
yes
Total exposure duration:
3 h
Post exposure observation period:
Not applicable
Test temperature:
21C
pH:
7.8-8.1
Nominal and measured concentrations:
1000
Details on test conditions:
TEST SYSTEM
- Test vessel: 500 Ml CONICAL FLASK
- Type (delete if not applicable): open / closed
- Material, size, headspace, fill volume:
- Aeration: CONTINUOUS
- Type of flow-through (e.g. peristaltic or proportional diluter): The mixture was aerated with compressed air via narrow bore glass tubes at a rate of approximately 0.5 - 1 litre per minute.
- Renewal rate of test solution (frequency/flow rate): not renewed
- No. of organisms per vessel:
- No. of vessels per concentration (replicates):3
- No. of vessels per control (replicates):3
- No. of vessels per vehicle control (replicates):3
- Biomass loading rate:

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: The test water used for both the range-finding and definitive studies was laboratory tap water dechlorinated by passage through an activated carbon filter (purite Series 500) and partly softened (Elga Nimbus 12480 Duplex Water Softener) giving water with a total hardness of approximately 100 mgll as CaC03.
- Total organic carbon:
- Particulate matter:
- Metals:
- Pesticides:
- Chlorine: 0.7 g NaCI, 0.4 g CaCI2.2H20 /L water
- Alkalinity: ca pH 7.5
- Ca/mg ratio: 2
- Conductivity:
- Culture medium different from test medium: same
- Intervals of water quality measurement:

OTHER TEST CONDITIONS
- Adjustment of pH:
- Photoperiod:
- Light intensity:

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :

TEST CONCENTRATIONS
- Spacing factor for test concentrations:
- Justification for using less concentrations than requested by guideline:
- Range finding study
- Test concentrations:
- Results used to determine the conditions for the definitive study:
Reference substance (positive control):
yes
Remarks:
3, 5-dichlorophenol
Key result
Duration:
3 h
Dose descriptor:
EC50
Effect conc.:
> 1 000 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
inhibition of total respiration
Remarks:
respiration rate
Key result
Duration:
3 h
Dose descriptor:
NOEC
Effect conc.:
1 000 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
inhibition of total respiration
Remarks:
respiration rate
Validity criteria fulfilled:
yes
Conclusions:
The effect of the test material on the respiration of activated sewage sludge microorganisms gave a 3-Hour ECso greater than 1000 mgll. Correspondingly the No Observed Effect Concentration (NOEC) was 1000 mgll.

Description of key information

Key value for chemical safety assessment

EC50 for microorganisms:
1 000 mg/L
EC10 or NOEC for microorganisms:
1 000 mg/L

Additional information

The EC50 and NOEC values refer to nominal loading levels resulting from water accomodated fraction (WAF). Water solubility of the substance is < 0.1 mg/l.